PROJECT SUMMARY Despite dramatic improvements in HIV-related morbidity and mortality3 since the introduction of combination antiretroviral therapy (ART), metabolic complications including insulin resistance (IR),4,5 adipose tissue changes,6-8 and cardiovascular disease (CVD)11,12 remain significant concerns in HIV-infected adults on ART. The association between ART and HIV-associated changes in body composition13 as well as IR has been documented.6-8,14 However, appreciable data is still lacking in potential biomarkers which may predict the development of poor metabolic outcomes. Metabolomics provides the ability to identify and quantify small molecules present in both disease and non- disease states, thereby distinguishing particular clusters of metabolites which could potentially serve as novel clinical biomarkers.18 Several non-HIV studies have shown a positive association of intermediary metabolites such as acylcarnitines (ACs) and branched chain amino acids (BCAAs) with IR,19-24 associations of BCAAs with obesity, and associations of phospholipids (PLs) and abnormal fatty acid synthesis with obesity.25-27 However, few studies have used targeted metabolomics to assess the association between these metabolites (ACs, PLs, BCAAs) and visceral adiposity or IR in HIV-infected adults29 ? particularly those on newer line ART - or explored whether these associations are different compared to an HIV-uninfected population. We will leverage the highly established research infrastructure within the AIDS Clinical Trials Group (ACTG) 5260s, Multicenter AIDS Cohort Study (MACS), and Women's Interagency HIV Study (WIHS) cohorts, utilizing repository specimens to study metabolomic signatures of central adiposity and IR in HIV-infected and -uninfected adults. ACTG A5257 (parent study of A5260s) was a randomized trial comparing atazanavir/ritonavir, darunavir/ritonavir and raltegravir-based ART. Well-characterized longitudinal data up to 96 weeks on the outcomes for this proposal have already been measured: central adiposity defined as visceral adiposity tissue (VAT) measured via serial CT scans in ACTG 5260s and waist circumference as well as IR measured using the Homeostatic Model Assessment ? IR (HOMA- IR)28 in all cohorts. MACS and WIHS are cohorts of HIV-infected and -uninfected men and women respectively where data and specimens from matched uninfected adults will be used. We will perform targeted metabolomics on longitudinally collected banked plasma specimens in ACTG 5260s to identify whether increased plasma ACs, PLs, AAs/BCAAs signatures predict increases in visceral adiposity and IR after initiation of newer line ART regimens. We will also perform targeted metabolomics on a cross-sectional sample of banked plasma specimens in ACTG 5260s, MACS, and WIHS to evaluate differences between HIV-infected and uninfected adults in plasma ACs, PLs, AAs/BCAAs and their relationship to central adiposity and IR. In addition to expanding our understanding of the pathogenesis and interplay between IR and the capacity of adipocytes to accumulate fat, results from our study will reveal novel biomarker signatures which may identify those at increased risk for the development of IR and central adiposity, thereby informing future prevention strategies against diabetes and CVD in HIV-infected individuals during an era of newer line ART.